FIG. 6.

Expression of the M. tuberculosis rpf-like genes in vitro as assessed by quantitative dilutional PCR. RNA was prepared from Erdman grown in 7H9 medium, and 1 μg of total RNA was reverse transcribed and serially diluted fivefold for use as template in PCRs to amplify each of the rpf-like genes. Primers, as described in Materials and Methods, were used to amplify portions of Rv0867c (A), Rv1009 (B), Rv1884c (C), Rv2389c (D), and Rv2450c (E). (A to E) Lanes 1 to 5 contain PCR products of serial dilutions of cDNA from day 4 (OD₆₀₀ = 0.125), lanes 6 to 10 are from day 6 (OD₆₀₀ = 0.574), lanes 11 to 15 are from day 8 (OD₆₀₀ = 1.408), and lanes 16 to 20 are from day 12 (OD₆₀₀ = 2.512). (F) PCR products of cDNAs from day 4 (lanes 1 and 5), day 6 (lanes 2 and 6), day 8 (lanes 3 and 7), and day 12 (lanes 4 and 8) using 16S rRNA primers (lanes 1 to 4, 9, and 10) and 23S rRNA primers (lanes 5 to 8, 11, and 12). Lanes 9 and 11 contain no template (negative controls), while lanes 10 and 12 contain Erdman genomic DNA as the template. Lanes 16 to 19 show that no PCR product was obtained from non-reverse-transcribed RNA from day 4 (lane16), day 6 (lane 17), day 8 (lane 18), or day 12 (lane 19) cultures, using primers to amplify Rv1884c. A 50-bp ladder (Invitrogen) is shown in the far left lane of each gel.